Synapses are specialized cell adhesions that are the fundamental functional units of the nervous system, but the extracellular signals that induce CNS synapse formation are poorly understood. By using highly purified populations of neurons and glia, we have recently observed that the number of synapses on retinal ganglion cells (RGCs), as well as other types of CNS neurons, in culture is enhanced 7-fold by soluble signals released by astrocytes. These data add to the growing evidence that extracellular signals powerfully regulate CNS synaptogenesis. In our preliminary studies, we have identified 2 proteins that appear to play a crucial role in astrocyte-enhanced synaptogenesis. First, we have found that thrombospondin (TSP), a large matrix-associated protein released by astrocytes in vitro and in vivo, is sufficient to induce the astrocytes, and is also sufficient to induce the formation of structural synapses in vitro, and is necessary for astrocytes to induce synaptogenesis in vitro. Second, we have found that the complement protein C1q is highly upregulated in neurons by astrocytes, and is also sufficient to induce the formation of structural synapses in vitro. Both proteins are localized to synapses throughout the developing brain. TSP and C1q are thus among the first few identified soluble proteins sufficient to trigger formation of structural synapses between CNS neurons in vitro. We propose to further investigate the molecular basis of TSP and C1q in inducing synaptogenesis in vitro and then use this knowledge to directly test the hypothesis that they also help to control synaptogenesis in the developing visual system. Understanding the mechanisms that regulate synaptogenesis and synaptic plasticity are crucial to understanding the neural basis of learning and memory, Alzheimer's disease, drug addiction, and epilepsy.